SAtBatSeTmReAntCoTf Hypothesis and Specific Aims Age is the single largest risk factor for the development of neoplasia. Investigation into how age contributes to increased cancer incidence has focused on accumulation of autonomous mutations within incipient cancer cells. While it is clear that these cell autonomous changes are integral to the transformation process, it has become evident that changes in the surrounding, ostensibly normal stroma collaborate in the process and may contribute to the age-dependent increase in cancer incidence. Indeed, normal fibroblasts within a tumor secrete factors that promote tumor cell growth. Like genetic mutations, senescent fibroblasts accumulate with age, and recent data suggests that they play a pivotal role in tumorigenicity. We hypothesize that as senescent fibroblasts increase within the stromal compartment they create a pro-tumorigenic environment that supports the growth and continued transformation of preneoplastic cells. To identify senescent fibroblast-derived factors that support preneoplastic cell growth, we carried out a nonbiased microarray analysis comparing young and senescent fibroblasts. We identified osteopontin (OPN) as a putative stromal factor capable of enhancing preneoplastic cell proliferation and thus hypothesize that it plays an important role in tumorigenesis. In support of this hypothesis, we found that OPN is expressed within the stromal compartment of benign human lesions of the skin and breast. While OPN expression has been correlated with tumor progression (33, 34), the role that stromal-derived OPN plays in early lesions has not been investigated. The goal of this proposal is to determine the molecular mechanisms by which OPN influences the early stages of the transformation process. To that end, the specific aims of this proposal are: Aim 1. Identify the receptor complex(es) engaged by fibroblast-derived OPN and the membrane proximal signaling molecules activated upon receptor binding Aim 2. Determine how fibroblast-derived OPN stimulates preneoplastic cell proliferation Aim 3. Determine how OPN expression is regulated upon activation of senescence Aim 4. Determine the impact of fibroblast-derived OPN on tumorigenesis